Means for inactivating pathogenic agents on surfaces, instruments and in contaminated fluids

ABSTRACT

The invention relates to ecologically-acceptable agent for treating pathogenic germs on surfaces, instruments and in fluids, comprising synergistic mixtures of aromatic hydroxybenzoic acids and phenols with a broad spectrum of action. The above is active against hydrophilically-sheathed and -unsheathed viruses as well as lipophilic bacteria and yeasts and is thus applicable in medicine, industry and commercial animal raising.

BACKGROUND OF THE INVENTION

Infections acquired by patients in hospitals and other medicalestablishments cause great damage to the community of insured and to thenational economy. These infections, called nosocomial diseases, havebeen attributed in the past, predominantly to bacteria.

SUMMARY OF THE INVENTION

On one hand, this has been due to the fact that in the absence ofadequate medical diagnostics, many diseases having a mycological orviral genesis have not been recognized. On the other hand, infectionscaused by viruses and fungi have increased as a result of moderntherapeutic measures and also as a result of travel and globalinterconnections; for instance, epidemics witnessed in more recent timesin European animal husbandry such as mouth-and-foot disease, Aujeszky'sdisease, and swine fever, without any exception had a viral genesis. Inhospitals, viral infections such as those due to the Norwalk-likeviruses, rotaviruses, and adenoviruses, but also fungal infectionsleading to systemic mycoses and secondary infections, are diagnosedincreasingly.

This new situation, and the new knowledge, have had the effect that inrecent years, prophylactic measures such as procedures of disinfectionmust be reconsidered and conceived in a new way. Thus, a number ofstandardizing authorities demand that apart from bacteria, disinfectionshould also extend to particularly resistant fungi (e.g., Aspergillusniger) and viruses (e.g., poliovirus and adenovirus).

Disinfectants that are broadly applicable and have sufficient viricidaleffectiveness are nowadays used in a very limited way only, the reasonbeing side effects of the agents. This is particularly true for thealdehyde-type active agents, e.g., formaldehyde, glutaric dialdehyde,succinic dialdehyde, or glyoxal and their derivatives giving offaldehydes.

Up to now these components were regarded as the classical vectors of abroad antimicrobial and antiviral effectiveness in disinfectantformulations.

Formaldehyde and glutaric dialdehyde, which are the agents mostuniversally applicable for fighting pathogenic agents owing to, amongstother reasons, the lack of technical problems in their application, havebeen classified as toxic and are suspected of being carcinogenic.Comparable characteristics are assumed to exist with the otheraldehydes.

This causes users considering the potential risks to largely do withoutaldehyde-based disinfectants.

Other active agents that are available are not effective, or are onlyeffective in a limited way, against unsheathed viruses and certain kindsof fungi, because of the particular resistance of these targets, or canonly be used in a restricted way because of their unfavorable chemicaland physical properties.

This holds true for the class of per compounds, for iodine, substancesgiving off chlorine, alcohols, cationic surfactants, amphotericsurfactants, phenols, bases, acids, and compounds giving off activeoxygen.

The peracids for instance have a very highly diversified spectrum ofantimicrobial action, but can be applied in a very limited way onlyowing to their extreme corrosivity. Considerable problems arise inaddition from the lack of stability of this compound class.

While looking for an adequate alternative it has now been foundsurprisingly that the gap that had arisen may be closed when usingcertain mixtures consisting of aromatic hydroxycarboxylic acids andphenols, not only on account of the microbicidal effectiveness but alsoon account of the favorable toxicological and ecotoxicologicalproperties and a favorable compatibility with materials.

Subject matter of the present invention are agents for the inactivationof pathogenic germs (bacteria, fungi, and viruses, sheathed andunsheathed) that can be applied to surfaces and instruments of all kindsas well as in contaminated fluids. The potential areas of applicationare the most diverse, e.g., in the context of hospitals, doctors'offices, production spaces of food industries and all the way to thestables of livestock breeders.

Additional features and advantages of the present invention aredescribed in, and will be apparent from, the following DetailedDescription of the Invention.

DETAILED DESCRIPTION OF THE INVENTION

A synergistic action between the components of the disinfectant mixturesaccording to the invention has been demonstrated, not only for theviricidal properties but also for the bactericidal and fungicidalproperties. The enhancement of bactericidal and fungicidal action wasall the more surprising inasmuch as for the phenols and aromatichydroxycarboxylic acids used, excellent antimicrobial effects hadalready been known for the individual components.

Another remarkable feature is the unusual breadth of the spectrum ofactivity, which can be seen from the fact that the inactivation ofhydrophilic Picorna viruses is just as reliable as the killing oflipophilic fungi.

The examples and tables reported in the following serve to explain thepresent invention and to prove the synergism between the synergistsaccording to claim 1.

According to F. C. Kull and P. C. Eisman, Applied Microbiology, 9,538-541 (1946), a synergism can be regarded as proven when a result ofF<1 is calculated with the following formula:F=QA/Qa+QB/Qb,

where the symbols have the meaning: F < 1 Synergism F = 1 Additiveeffect F > 1 Antagonism Qa = quantity of A alone to end point Qb =quantity of B alone to end point QA = quantity of A in the mixture withB QB = quantity of B in the mixture with A.

EXAMPLES Example No. 1

Alkyl aryl sulfonate Na 12.0 parts by weight Butyl monoglycol sulfonateNa 5.0 4-Chloro-3-methylphenol 15.0 Phosphonobutanetricarboxylic acid1.5 2-Propyl alcohol 30.0 Water deionized 36.5

Example No. 2

Alkyl aryl sulfonate Na 12.0 parts by weight Butyl monoglycol sulfonateNa 5.0 2-Hydroxybenzoic acid 6.0 Phosphonobutanetricarboxylic acid 1.52-Propyl alcohol 30.0 Water deionized 45.5

Example No. 3

Alkyl aryl sulfonate Na 12.0 parts by weight Butyl monoglycol sulfonateNa 5.0 4-Chloro-3-methylphenol 15.0 2-Hydroxybenzoic acid 6.0 2-Propylalcohol 30.0 Phosphonobutanetricarboxylic acid 1.5 Water deionized 30.5

Example No. 4

Alkyl sulfonate Na 10.0 parts by weight Cumenesulfonate Na 3.02-Phenylphenol 15.0 β-Resorcinolic acid 7.0 Formic acid 5.0 2-Propylalcohol 33.0 Water deionized 27.0

Formulation Examples Nos. 1 to 3 were used to prove the synergisticeffect concerning viricidal properies with the combinations according tothe invention.

The unsheathed hydrophilic Picorna virus Polio Sabin LSc-2ab served asthe test criterion. In the testing procedure and method, the rules ofthe preliminary European standard WI 216026 (phase 2; step 1) werefollowed.

Experimental Conditions: Temperature 10° C. ± 1° C. Contact time 30 min± 10 s Protein load 3 g bovine serum albumin and 10 g yeast extract perliter

Unless pointed out otherwise, all numbers reported in the Table signifythe infection titer (log ID₅₀, ml⁻¹) after a 30-min period of action; -means no longer detectable. TABLE 1 (Polio Sabin) Concentrations (%)Sample Control 2% 3% 4% 5% 6% 7% Example No. 1 7.7 6.9 6.3 5.8 4.3 3.93.0 Example No. 2 7.3 6.2 5.5 4.3 3.8 2.8 2.3 Example No. 3 7.8 3.6 2.1— — — —

A result can be interpreted as sufficiently effective when the infectiontiter is reduced by four logarithmic steps, ie, when a 99.99% reductionof infectivity has been attained.

It follows from Table 1 that in Example 1, the phenol component waseffective in an applied concentration of 7%, while in Example 2, thehydroxybenzoic acid had a sufficiently strong effect at a concentrationof 6%. The mixture of the two components used in Formulation example No.3 revealed a sufficiently strong inactivation of the poliovirus, alreadyat a concentration of 2%.

Substituting these results into the formula of Kull and Eisman, we find:F=2×0.06/6×0.06+2×0.15/7×0.15=0.62.

The numerical value of 0.62 thus yields unambiguous proof for thepresence of a synergistic effect.

(Substituting the percent quantities of the active agents from theformulation examples into the equation produces a constant factor of 1,and hence is superfluous for the calculation.)

A synergistic effect in fungicidal effectiveness could be demonstratedin the instance of the particularly resistant Aspergillus niger.

The tests were conducted with the quantitative suspension test accordingto DIN EN 1650 (phase 2; step 1).

Experimental Conditions: DIN EN 1650

A germ count reduction by four logarithmic steps constitutes therequired proof of effectiveness.

The test results are reported in Table 2. The numerical values giventhere are the logarithms (log₁₀) of the reduced germ counts; thedifference between these counts and the original germ count yields thereduction factor. TABLE 2 (Asp. niger) Test solution log Concentrations(%) Examples (germ count/ml) 0.25% 0.5% 1% 2% 1 7.67 4.9 3.8 3.1 2 7.675.2 4.2 2.8 3 7.67 2.1

The above result of calculation from the data of Table 2 demonstratesthat here, too, a synergistic effect of the mixture of active agents ispresent.

The proof for synergistically effective properties against bacteria wasobtained in a quantitative suspension test according to DIN EN 1276(phase 2; step 1) with a Gram-positive and a Gram-negative testorganism.

Experimental conditions: Period of action: 20 min at 20° C. Proteinload: 3.0 g bovine serum albumin per liter.

Table 3 shows the results obtained with Escherichia coli. TABLE 3 (E.coli) Concentrations (%) Examples Log (germ count/ml) 0.25% 0.5% 1% 2% 18.49 4.63 4.1 2 8.49 5.15 4.9 2.9 3 8.49 3.16Table 4 shows the results obtained with Staphylococcus aureus.

Experimental conditions: DIN EN 1276 TABLE 4 (Staph. aureus)Concentrations (%) Examples Log (germ count/ml) 0.25% 0.5% 1% 2% 1 8.45.9 4.8 3.3 2 8.4 6.3 5.5 4.9 2.8 3 8.4 5.1 3.17

A germ count reduction by four logarithmic steps represented therequired proof of effectiveness.F=0.5/1+0.5/2=0.75.

The microbiological results needed to prove synergism could be obtainedin all the tests, which demonstrates that the formulations according toclaim 1 of the invention are synergistically effective against bacteria,fungi, and viruses.

The formulation according to Example No. 4 and the Mycobacterium aviumAv 56 served to obtain proof of a tuberculocidal effect in the germcarrier test. The test conditions matched the provisions of the GermanVeterinary-Medical Society for the Area of Animal Husbandry (2^(nd)edition 1998).

Germ carrier: sterilized limewood pieces (height 3 mm, length 10 mm,width 10 mm). TABLE 5 (Mycobacterium avium) Example No. 4 Period ofaction [min] Concentration [%] 30 60 120 180 240 2 + + + + + 4 + + − − −5 + + − − − 6 + + − − − Formalin 3% + + − − − Growth control + + + + +

The test result presented in Table 5 shows that a 4% solution ofFormulation example No. 4 after a period of action of 120 min yields thesame effect as a 3% solution of Formalin. According to DAB 10 (GermanPharmacopoeia 10^(th) edition), Formalin contains 35 to 37% formaldehydein water and 10% methanol, which corresponds to an effectiveconcentration of about 1.1% aldehyde.

In the formulation according to Example No. 4, 15%+7% of an effectivemixture of substances are present, of which 4% are employed, whichcorresponds to an effective concentration of the active agent of only0.88%.

Formalin is the generally recognized reference and scale in germ carriertests on limewood, since the relatively small aldehyde molecule willparticularly well penetrate into the fissured and disintegrated fiberstructure of the limewood carrier, and act there.

The result of the tuberculocidal effectiveness test underlines, justlike the other results, that the present invention meets all conditionsfor being able to replace aldehyde-based disinfectant formulations.

It should be understood that various changes and modifications to thepresently preferred embodiments described herein will be apparent tothose skilled in the art. Such changes and modifications can be madewithout departing from the spirit and scope of the present invention andwithout diminishing its intended advantages. It is therefore intendedthat such changes and modifications be covered by the appended claims.

1. A method for the control and inactivation of pathogenic germs onsurfaces and instruments of medical and technical establishmentscomprising the step of using an effective microbicidal and antiviralcombination of at least one acid selected from the group consisting ofaromatic monohydroxycarboxylic acids, dihydroxybenzoic acids, andtrihydroxybenzoic acids individually or mixed, and phenols incombination with a surfactant selected from the group consisting ofalkyl sulfonates, alkyl aryl sulfonic acid, alkyl aryl sulfonates, alkylaryl ether sulfates with 1 to 3 EO groups, alkyl ether sulfates with 1to 3 EP groups, their sodium, potassium, and ammonium salts with primaryor branched chains having a length of C₈ to C₁₈ as anionic surfactantand alkyl polyethyleneglycol ethers with 3 to 11 EO groups as nonionicsurfactant, individually or mixed.
 2. The method according to claim 1,wherein the disinfectant comprises at least one salt selected from thegroup consisting of butyl monoglycol sulfate, cumenesulfonate,toluenesulfonate, xylenesulfonate as sodium, potassium, or ammoniumsalt, and combinations there of, as a hydrotropic agent, and aliphaticalcohols or glycols having a chain length of C₂ to C₁₂, individually oras a mixture, as a solvent, and aliphatic carboxylic acids orhydroxycarboxylic acids having a chain length of C₁ to C₆, individuallyor as a mixture, as pH regulators.
 3. The method according to claim 1the weight ratio of the hydroxybenzoic acids (A) to the phenols (B) isbetween 1:9 and 9:1, and their sum is between 5 and 40% by weight,referring to the total weight of the concentrated disinfectant formula.4. The method according to claim 1 wherein the weight ratio of the alkylsulfonates and/or alkyl aryl sulfates and/or ether sulfates and theirsalts (C) to the acids and phenols (A+B), C: (B+A), is between 1:9 and9:1, and their sum is between 10 and 60%, referring to the total weightof the concentrated disinfectant formula.
 5. The method Use according toclaim 2 wherein the weight ratio of the hydrotropic agents and theirsalts, individually or in their mixture, is between 5 and 40% by weight,referring to the total weight of the concentrated disinfectant formula.6. The method according to claim 2 wherein the weight ratio of thealcohols, individually or in their mixture, is between 5 and 60% byweight, referring to the total weight of the concentrated disinfectantformula.
 7. The method according to claim 1 wherein the disinfectantcontains between 1 and 8% by weight of at least one sequestering agentselected from the group consisting of aminoacetic acids and phosphonicacids and their derivatives.
 8. The method according to claim 1 whereinthe combination is in an aqueous, dilute solution containing between 0.5and 10% by weight of the concentrated disinfectant formula.
 9. Themethod according to claim 1 wherein the phenols are selected from thegroup consisting of 2-isopropyl-5-methylphenol, 2-, 3-, or4-methylphenol, hexylresorcinol, 2-phenylphenol, 2-methoxyphenol,3-methyl-4-chlorophenol, 3,5-dimethyl-4-chlorophenol,2-benzyl-4-chlorophenol individually or mixed.
 10. The method accordingto claim 1 wherein the aromatic monohydroxycarboxylic acid is selectedfrom the group consisting of 2-; 3-; 4-hydroxybenzoic acid.
 11. Themethod according to claim 1 wherein the dihydroxybenzoic acids areselected from the group consisting of 2,3-; 2,4-; 2,5-; 2,6-; 3,4-; and3,5-dihydroxybenzoic acid.
 12. The method according to claim 1 whereinthe trihydroxybenzoic acid is selected from the group consisting of2,3,4-trihydroxybenzoic acid, 2,4,6-trihydroxybenzoic acid,3,4,5-trihydroxybenzoic acid.
 13. A method of preparing a product foruse as a disinfectant for the control and inactivation of pathogenicgerms comprising the steps of producing an effective mixture of mono,di, and trihydroxybenzoic acids and phenols and anionic and nonionicsurfactants as wetting agents by mixing an effective microbicidal andantiviral combinations of at least one acid selected from the groupconsisting of aromatic monohydroxycarboxylic acids, dihydroxybenzoicacids, and trihydroxybenzoic acids, and phenols in combination with asurfactant selected from the group consisting of alkyl sulfonates, alkylaryl sulfonic acid, alkyl aryl sulfonates, alkyl aryl ether sulfateswith 1 to 3 EO groups, alkyl ether sulfates with 1 to 3 EP groups, theirsodium, potassium, and ammonium salts with primary or branched chainshaving a length of C₈ to C₁₈ as anionic surfactant and alkylpolyethyleneglycol ethers with 3 to 11 EO groups as nonionic surfactant.14. The method according to claim 13, wherein the disinfectant comprisesa salt selected from the group consisting of butyl monoglycol sulfate,cumenesulfonate, toluenesulfonate, xylenesulfonate as sodium, potassium,or ammonium salt; at least one aliphatic alcohols or glycols having achain length of C₂ to C₁₂; and at least one aliphatic carboxylic acidsor hydroxycarboxylic acids having a chain length of C₁ to C₆, as pHregulators.
 15. The method according to claim 13 wherein the weightratio of the hydroxybenzoic acids (A) to the phenols (B) is between 1:9and 9:1.
 16. The method according to claim 13 wherein the weight ratioof the alkyl sulfonates and/or alkyl aryl sulfates and/or ether sulfatesand their salts (C) to the acids and phenols (A+B), C:(B+A), is between1:9 and 9:1.
 17. The method according to claim 1 wherein thedisinfectant contains between 1 and 8% by weight of at least onesequestering agent.
 18. The method according to claim 13 comprising thestep of preparing an aqueous, dilute solution containing between 0.5 and10% by weight of the concentrated disinfectant formula.
 19. The methodaccording to claim 13 wherein the phenols are selected from the groupconsisting of 2-isopropyl-5-methylphenol, 2-, 3-, or 4-methylphenol,hexylresorcinol, 2-phenylphenol, 2-methoxyphenol,3-methyl-4-chlorophenol, 3,5-dimethyl-4-chlorophenol,2-benzyl-4-chlorophenol individually or mixed.
 20. The method accordingto claim 13 wherein the aromatic monohydroxycarboxylic acid is selectedfrom the group consisting of 2-; 3-; 4-hydroxybenzoic acid.
 21. Themethod according to claim 13 wherein the dihydroxybenzoic acids areselected from the group consisting of 2,3-; 2,4-; 2,5-; 2,6-; 3,4-; and3,5-dihydroxybenzoic acid.
 22. The method according to claim 13 whereinthe trihydroxybenzoic acid is selected from the group consisting of2,3,4-trihydroxybenzoic acid, 2,4,6-trihydroxybenzoic acid,3,4,5-trihydroxybenzoic acid.